Neuroblastoma is the most common extracranial tumor of childhood, and 60% of patients have aggressive disease from which only 20-40% survive. Their long term goal is to gain a better molecular biologic understanding of aggressive tumors so that more effective therapies can be developed. Their previous research suggests that the proto-oncogene MYCN, the neurotrophin receptors TrkA and p75TNR, and the neurotrophin Nerve Growth Factor (NGF) have important roles in determining the behavior of neuroblastomas. Hypothesis. Aggressive neuroblastomas escape the growth inhibitory, differentiating, and apoptotic influences of TrkA and p75TNR 1) by not expressing these receptors; 2) by altering signal transduction downstream from these receptors; and/or 3) by creating a NGF/p75NTR autocrine, anti-apoptotic loop. Specific Aims. 1) Define relationships between expression of TrkA, p75TNR , NGF, MYCN, MYCN gene status, histopathology, and clinical behavior of neuroblastomas. 2) Determine with retroviral gene transduction models if p75TNR can induce apoptosis and if a NGF/p75TNR autocrine loop enhances survival of neuroblastoma cells. 3) Determine if a naturally occurring NGF/p75TNR autocrine loop enhances survival of neuroblastoma cells. Research Design. RT-PCR quantitation of TrkA, p75TNR, NGF, and MYCN mRNA will be completed for 360 neuroblastomas. Histopathology will characterize tumors morphologically and identify proliferating and apoptotic cells and cells expressing p75TNR, TrkA, and NGF. Clinically important subsets will be defined. To assess the role of p75TNR in apoptosis, cell lines will be transduced with p75TNR in the presence of NGF, and then their dependence upon NGF for survival will be determined. Studies of p75TNR mediated signaling will emphasize roles of TrkA co-expression and of sphingomyelin/ceramide and caspases in apoptosis. To demonstrate that p75TNR and NGF can form an autocrine loop, neuroblastoma cell lines that have been transduced with p75TNR and shown to undergo apoptosis upon NGF withdrawal will be co-transduced with NGF cDNA. Approximately 60 low passage cell lines established at diagnosis will be screened to identify those expressing NGF and p75TNR (RT-PCR) and retaining sensitivity to ceramide. These will be tested with p75TNR and TrkA specific antagonists and an Ig-TrkA fusion protein to determine if they have a natural NGF/p75TNR autocrine loop that prevents apoptosis. Methods. These include quantitative RT-PCR, gene transduction, assessing clonogenic growth in microwells with digital image microscopy, immunostaining for TrkA and p75TNR selective agonist and antagonist molecules and with an Ig-TrkA fusion protein. We anticipate that this research will contribute to the understanding of neuroblastoma and ultimately to therapeutic modalities that are more effective and specific.